UniTaq Prime
Multiplex PCR Designs That Work
First Time, Every Time
UniTaq Prime is a comprehensive, easy-to-use set of software tools for multiplex PCR
Why UniTaq
A single set of oligos always works — eliminating the need for redesigns and backup assays: saving time and money. This differentiates our tools from academic (1) and commercial (2) PCR design solutions that describe “assay redesign” and wet lab testing of several candidate assays as an integral part of PCR R&D. Your R&D team will have a multiplex PCR test candidate ready for validation in under a month (when oligos arrive), compared to the typical 3+ months.
Our clients report achieving “the highest RFI ever seen in the Lab.” High RFI (Relative Fluorescent Intensity) increases test robustness: reducing invalid results and delivering the best possible LOD (limit of detection). Fewer invalid results, in turn, decrease the cost of customer support for IVD tests and the need for retesting for LDTs.
References
1. Bustin S, Huggett J. qPCR primer design revisited. Biomolecular Detection and Quantification. 2017 Dec;14:19–28.
2. SantaLucia J, Sozhamannan S, Gans JD, Koehler JW, Soong R, Lin NJ, et al. Appendix Q: Recommendations for Developing Molecular Assays for Microbial Pathogen Detection Using Modern In Silico Approaches. J AOAC Int. 2020 July 1;103(4):882–99.
Speed up R&D for qPCR and dPCR
Multiplex PCR test candidate in <1 month vs. 3+ months: no “redesign”
Our customers: “the strongest qPCR signal we have seen in our lab”
Fewer invalid
test results
Spend less on customer support
Shorten test turnaround time and lower costs by decreasing repeat testing
Respiratory panel performance
As a proof-of-performance example we designed and tested respiratory panel qPCR 5-plex: infA, infB, RSV, SARS-CoV2 and RPP30 mRNA IPC (Internal Positive Control). A single set of oligos was synthesized; all 15 viral oligos were pooled by oligo vendor -> we skipped single-plex testing and oligo concentration optimization. The figure shows excellent qPCR performance for viral serial dilutions: all four viruses show strong RFI signal and LOD of <10 copies/well. The RFI decreases only slightly at high Ct values with <10 target molecules per well.
Experiment details
- 20 µL reactions: no human RNA and + human RNA (10 ng/well): growth curves essentially overlap.
- All 15 viral oligos were pooled by IDT; final oligo concentrations: 500/200 nM primer / probe; RPP30 IPC at 250/100 nM TYE705 dye.
- Thermo TaqPath 1-step RT-PCR MMx used; data are similar in NEB LUNA MMx;
- PCR efficiency is ~100% in both MMxes.
- The lowest titer is <10 copies/well; next serial dilution had no qPCR signal.
UniTaq Prime: software feature highlights
1. Complete solution
UniTaq Prime delivers a fully integrated solution for multiplex PCR design. Two software tools download sequences from NCBI: ncbi_genome — viral genomes; ncbi_gene — gene sequences based on NCBI taxonomy IDs. Both tools output multi-fasta files, which serve as input for the design tool. that generates candidate assays. The latter are scored for >25 parameters that affect qPCR RFI. High scoring assays are used as input for the multiplex tool.
2. Web front-end
All UniTaq Prime tools feature a web-based front-end, making them easy to use by scientists. All tools generates multi-worksheet Excel files, enabling you to review results, filter, sort and select assay [and multiplex] candidates for synthesis and wet lab validation.
3. UniTaq Prime: details
UniTaq Prime software tools can be licensed to fit your needs: infectious disease qPCR or Human targets: SNP/mutations, gDNA/CNV, GEx and PCR enrichment. We also offer professional services: design PCR multiplexes based on your requirements.
Our focus is on the three segments of Molecular Diagnostics (MDx) market: 1. MDx vendors developing IVD tests; 2. Diagnostics labs running LDTs; 3. PCR reagent vendors.
We describe in more detail multiple factors that affect qPCR performance. When >25 factors are accounted for – the multiplex PCR always works and qPCR signal (RFI) is strong: a single set of oligos is sufficient to start your validation work.
4. Runs on the cloud
UniTaq Prime runs on Google cloud (GCP), a leading cloud provider, ensuring enterprise-grade security and scalability. We leverage Galaxy – a widely adopted academic software (3) – to deploy all tools online. Galaxy workflows allow several tools to run sequentially with a single click.
Reference
3. Jalili V, Afgan E, Gu Q, Clements D, Blankenberg D, Goecks J, et al. The Galaxy platform for accessible, reproducible and collaborative biomedical analyses: 2020 update. Nucleic Acids Research. 2020 July 2;48(W1): W395–402.
Interested in our Multiplex PCR Designs?
